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- PDB-4jy6: Crystal structure of human Fab PGT123, a broadly reactive and pot... -

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Basic information

Entry
Database: PDB / ID: 4jy6
TitleCrystal structure of human Fab PGT123, a broadly reactive and potent HIV-1 neutralizing antibody
Components
  • PGT123 heavy chain
  • PGT123 light chain
KeywordsIMMUNE SYSTEM / Broadly neutralizing antibody against HIV-1 / HIV-1 Env gp120 subunit
Function / homologyImmunoglobulins / Immunoglobulin-like / Sandwich / Mainly Beta
Function and homology information
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.5 Å
AuthorsJulien, J.-P. / Diwanji, D.C. / Burton, D.R. / Wilson, I.A.
CitationJournal: PLoS Pathog / Year: 2013
Title: Broadly neutralizing antibody PGT121 allosterically modulates CD4 binding via recognition of the HIV-1 gp120 V3 base and multiple surrounding glycans.
Authors: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael ...Authors: Jean-Philippe Julien / Devin Sok / Reza Khayat / Jeong Hyun Lee / Katie J Doores / Laura M Walker / Alejandra Ramos / Devan C Diwanji / Robert Pejchal / Albert Cupo / Umesh Katpally / Rafael S Depetris / Robyn L Stanfield / Ryan McBride / Andre J Marozsan / James C Paulson / Rogier W Sanders / John P Moore / Dennis R Burton / Pascal Poignard / Andrew B Ward / Ian A Wilson /
Abstract: New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, ...New broad and potent neutralizing HIV-1 antibodies have recently been described that are largely dependent on the gp120 N332 glycan for Env recognition. Members of the PGT121 family of antibodies, isolated from an African donor, neutralize ∼70% of circulating isolates with a median IC50 less than 0.05 µg ml(-1). Here, we show that three family members, PGT121, PGT122 and PGT123, have very similar crystal structures. A long 24-residue HCDR3 divides the antibody binding site into two functional surfaces, consisting of an open face, formed by the heavy chain CDRs, and an elongated face, formed by LCDR1, LCDR3 and the tip of the HCDR3. Alanine scanning mutagenesis of the antibody paratope reveals a crucial role in neutralization for residues on the elongated face, whereas the open face, which accommodates a complex biantennary glycan in the PGT121 structure, appears to play a more secondary role. Negative-stain EM reconstructions of an engineered recombinant Env gp140 trimer (SOSIP.664) reveal that PGT122 interacts with the gp120 outer domain at a more vertical angle with respect to the top surface of the spike than the previously characterized antibody PGT128, which is also dependent on the N332 glycan. We then used ITC and FACS to demonstrate that the PGT121 antibodies inhibit CD4 binding to gp120 despite the epitope being distal from the CD4 binding site. Together, these structural, functional and biophysical results suggest that the PGT121 antibodies may interfere with Env receptor engagement by an allosteric mechanism in which key structural elements, such as the V3 base, the N332 oligomannose glycan and surrounding glycans, including a putative V1/V2 complex biantennary glycan, are conformationally constrained.
History
DepositionMar 29, 2013Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 8, 2013Provider: repository / Type: Initial release
Revision 1.1May 15, 2013Group: Database references
Revision 1.2Jun 19, 2013Group: Database references
Revision 1.3Sep 20, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_struct_conn_angle / struct_conn / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: PGT123 light chain
B: PGT123 heavy chain
C: PGT123 light chain
D: PGT123 heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)97,06423
Polymers95,7954
Non-polymers1,26919
Water2,972165
1
A: PGT123 light chain
B: PGT123 heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,77515
Polymers47,8982
Non-polymers87713
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area5290 Å2
ΔGint-320 kcal/mol
Surface area22530 Å2
MethodPISA
2
C: PGT123 light chain
D: PGT123 heavy chain
hetero molecules


Theoretical massNumber of molelcules
Total (without water)48,2908
Polymers47,8982
Non-polymers3926
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4410 Å2
ΔGint-153 kcal/mol
Surface area22380 Å2
MethodPISA
Unit cell
Length a, b, c (Å)84.890, 80.320, 90.430
Angle α, β, γ (deg.)90.00, 93.85, 90.00
Int Tables number4
Space group name H-MP1211

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Components

#1: Antibody PGT123 light chain


Mass: 22728.102 Da / Num. of mol.: 2 / Fragment: Fab
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / Strain (production host): Sf9
#2: Antibody PGT123 heavy chain


Mass: 25169.400 Da / Num. of mol.: 2 / Fragment: Fab
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / Strain (production host): Sf9
#3: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 18 / Source method: obtained synthetically / Formula: Zn
#4: Chemical ChemComp-GOL / GLYCEROL / GLYCERIN / PROPANE-1,2,3-TRIOL


Mass: 92.094 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C3H8O3
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 165 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.21 Å3/Da / Density % sol: 61.69 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 6.5
Details: 0.16 M zinc acetate, 0.08 M sodium cacodylate, pH 6.5, 20% v/v glycerol, 20% w/v PEG8000, VAPOR DIFFUSION, SITTING DROP, temperature 293K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-B / Wavelength: 1.033 Å
DetectorType: MARMOSAIC 300 mm CCD / Detector: CCD / Date: Nov 11, 2010
RadiationMonochromator: Double crystal cryo-cooled Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.033 Å / Relative weight: 1
ReflectionResolution: 2.5→30 Å / Num. all: 42191 / Num. obs: 42069 / % possible obs: 99.7 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 3.7 % / Rsym value: 0.118 / Net I/σ(I): 9.2
Reflection shellResolution: 2.5→2.6 Å / Redundancy: 3.8 % / Mean I/σ(I) obs: 2.3 / Num. unique all: 4187 / Rsym value: 0.51 / % possible all: 99.9

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Processing

Software
NameVersionClassification
Blu-Icedata collection
PHASERphasing
PHENIX(phenix.refine: 1.7.2_869)refinement
XDSdata reduction
XDSdata scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 3MUH

3muh


Resolution: 2.5→29.141 Å / SU ML: 0.84 / σ(F): 0 / Phase error: 29.54 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.2652 2086 5 %RANDOM
Rwork0.2069 ---
all0.2099 42191 --
obs0.2099 41693 98.93 %-
Solvent computationShrinkage radii: 0.98 Å / VDW probe radii: 1.2 Å / Solvent model: FLAT BULK SOLVENT MODEL / Bsol: 57.433 Å2 / ksol: 0.35 e/Å3
Displacement parameters
Baniso -1Baniso -2Baniso -3
1-9.5963 Å20 Å28.2532 Å2
2---6.9369 Å2-0 Å2
3----2.6593 Å2
Refinement stepCycle: LAST / Resolution: 2.5→29.141 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6622 0 24 165 6811
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0116808
X-RAY DIFFRACTIONf_angle_d1.2269286
X-RAY DIFFRACTIONf_dihedral_angle_d16.6022419
X-RAY DIFFRACTIONf_chiral_restr0.0791053
X-RAY DIFFRACTIONf_plane_restr0.0071186
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.5-2.55820.42361360.33722555X-RAY DIFFRACTION97
2.5582-2.62210.40451480.32752580X-RAY DIFFRACTION98
2.6221-2.6930.37731250.30512625X-RAY DIFFRACTION98
2.693-2.77220.31681410.27682637X-RAY DIFFRACTION99
2.7722-2.86160.34281140.25722630X-RAY DIFFRACTION98
2.8616-2.96370.32231410.24922607X-RAY DIFFRACTION99
2.9637-3.08230.29361410.23562667X-RAY DIFFRACTION99
3.0823-3.22240.31511380.23092606X-RAY DIFFRACTION99
3.2224-3.3920.2671250.21642660X-RAY DIFFRACTION99
3.392-3.60420.25211530.19872649X-RAY DIFFRACTION100
3.6042-3.88190.27281330.19212671X-RAY DIFFRACTION99
3.8819-4.27150.24911610.17932642X-RAY DIFFRACTION100
4.2715-4.8870.20021430.14412656X-RAY DIFFRACTION100
4.887-6.14760.21551480.1692678X-RAY DIFFRACTION99
6.1476-29.14260.22291390.19362744X-RAY DIFFRACTION99
Refinement TLS params.Method: refined / Origin x: -13.5205 Å / Origin y: 8.8467 Å / Origin z: 24.9009 Å
111213212223313233
T0.3153 Å2-0.0124 Å20.073 Å2-0.165 Å2-0.006 Å2--0.2077 Å2
L0.2723 °2-0.0099 °2-0.0916 °2-0.2127 °2-0.0726 °2--0.3184 °2
S-0.0317 Å °0.0476 Å °-0.0482 Å °-0.0795 Å °0.0069 Å °-0.0216 Å °0.0611 Å °-0.0081 Å °0.0241 Å °
Refinement TLS groupSelection details: all

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